Environmental Enrichment: effects on the visual system of an animal model of Retinitis pigmentosa (RP)

La Retinite Pigmentosa (RP) comprende una famiglia di degenerazioni ereditarie eterogenee (Kennan et al. 2005) (Paskowitz et al. 2006) caratterizzate dall’iniziale morte apoptotica dei bastoncelli, con sviluppo di insufficiente capacità visiva in condizioni di scarsa luminosità e progressivo restringimento concentrico del campo visivo, cui segue la degenerazione dei coni che comporta il declino dell’acuità visiva, fino alla cecità. Sebbene la causa primaria della RP sia tipicamente un difetto di un gene specifico dei bastoncelli, la degenerazione di queste cellule attiva una cascata di eventi che portano sia alla morte secondaria dei coni che al cosiddetto “remodeling” progressivo dei neuroni della retina interna (Marc et al. 2003). Nella patologia umana come nei suoi modelli animali, questi eventi seguono un pattern spazialmente irregolare sulla superficie retinica, rendendo difficile un’analisi quantitativa della sopravvivenza e della morte cellulare. Lo scopo principale di questa tesi è quello di far luce sugli effetti precoci e tardivi dell’applicazione del paradigma sperimentale dell’Arricchimento Ambientale (AA) sulla degenerazione dei bastoncelli, sulla sopravvivenza dei coni e sulla morfologia dei neuroni interni della retina del mutante rd10, un modello murino di RP caratterizzato dalla progressiva degenerazione dei fotorecettori retinici secondo un pattern simile a quello che avviene nell’uomo affetto da analoghe mutazioni. L’AA è un paradigma di stimolazione motoria, sensoriale e sociale ampiamente usato come strategia non invasiva di neuroprotezione in diversi modelli di patologie neurologiche, incluse la malattia di Alzheimer e la malattia di Huntington. In questa tesi vengono riportati per la prima volta i risultati dell’esposizione prolungata di topi rd10 ad AA. Attraverso studi di microscopia del tessuto retinico, registrazioni elettroretinografiche, test comportamentali e analisi molecolari, viene mostrato che l’AA dalla nascita è in grado di preservare in modo considerevole la morfologia, la fisiologia dei fotorecettori e la percezione visiva negli animali rd10. Studi preliminari suggeriscono che l’effetto protettivo dell’AA possa essere mediato da fattori neurotrofici, tra i quali il CNTF. Questi risultati possono incoraggiare lo sviluppo di nuove strategie applicative in cui la stimolazione ambientale, da sola o in combinazione con opportuni trattamenti farmacologici, può promuovere la conservazione e l’integrità funzionale dei neuroni retinici

Tackling Lateral Variability Using Surface Waves: A Tomography-Like Approach

Lateral velocity variations in the near-surface reflect the presence of buried geological or anthropic structures, and their identification is of interest for many fields of application. Surface wave tomography (SWT) is a powerful technique for detecting both smooth and sharp lateral velocity variations at very different scales. A surface-wave inversion scheme derived from SWT is here applied to a 2-D active seismic dataset to characterize the shape of an urban waste deposit in an old landfill, located 15 km South of Vienna (Austria). First, the tomography-derived inverse problem for the 2-D case is defined: under the assumption of straight rays at the surface connecting sources and receivers, the forward problem for one frequency reduces to a linear relationship between observed phase differences at adjacent receivers and wavenumbers (from which phase velocities are straightforwardly derived). A norm damping regularization constraint is applied to ensure a smooth solution in space: the choice of the damping parameter is made through a minimization process, by which only phase variations of the order of the average wavelength are modelled. The inverse problem is solved for each frequency with a weighted least-squares approach, to take into account the data error variances. An independent multi-offset phase analysis (MOPA) is performed using the same dataset, for comparison: pseudo-sections from the tomography-derived linear inversion and MOPA are very consistent, with the former giving a more continuous result both in space and frequency and less artefacts. Local dispersion curves are finally depth inverted and a quasi-2-D shear wave velocity section is retrieved: we identify a well-defined low velocity zone and interpret it as the urban waste deposit body. Results are consistent with both electrical and electromagnetic measurements acquired on the same line

Intranasal rapamycin ameliorates Alzheimer-like cognitive decline in a mouse model of Down syndrome

Background: Down syndrome (DS) individuals, by the age of 40s, are at increased risk to develop Alzheimer-like dementia, with deposition in brain of senile plaques and neurofibrillary tangles. Our laboratory recently demonstrated the disturbance of PI3K/AKT/mTOR axis in DS brain, prior and after the development of Alzheimer Disease (AD). The aberrant modulation of the mTOR signalling in DS and AD age-related cognitive decline affects crucial neuronal pathways, including insulin signaling and autophagy, involved in pathology onset and progression. Within this context, the therapeutic use of mTOR-inhibitors may prevent/attenuate the neurodegenerative phenomena. By our work we aimed to rescue mTOR signalling in DS mice by a novel rapamycin intranasal administration protocol (InRapa) that maximizes brain delivery and reduce systemic side effects. Methods: Ts65Dn mice were administered with InRapa for 12 weeks, starting at 6 months of age demonstrating, at the end of the treatment by radial arms maze and novel object recognition testing, rescued cognition. Results: The analysis of mTOR signalling, after InRapa, demonstrated in Ts65Dn mice hippocampus the inhibition of mTOR (reduced to physiological levels), which led, through the rescue of autophagy and insulin signalling, to reduced APP levels, APP processing and APP metabolites production, as well as, to reduced tau hyperphosphorylation. In addition, a reduction of oxidative stress markers was also observed. Discussion: These findings demonstrate that chronic InRapa administration is able to exert a neuroprotective effect on Ts65Dn hippocampus by reducing AD pathological hallmarks and by restoring protein homeostasis, thus ultimately resulting in improved cognition. Results are discussed in term of a potential novel targeted therapeutic approach to reduce cognitive decline and AD-like neuropathology in DS individuals

Frequency-Domain Electromagnetic Mapping of an Abandoned Waste Disposal Site: A Case in Sardinia (Italy)

For decades, bad practices inmunicipal and industrial wastemanagement have had negative environmental impacts, generating high health risks for people and the environment. The use of badly designed, not engineered, and not well-operated landfills has, around the world, produced a large number of potentially contaminated sites, for which there are urgent needs to assess the actual risk and to proceed, in case, with reclamation activities. One of these sites, an abandoned waste disposal site located near a Site of Community Importance on the central-eastern coast of Sardinia (Italy), is the subject of the case history described in this work. As a part of a multi-method geophysical characterisation, a frequency-domain electromagnetic (FDEM) mapping survey was carried out with the specific aim of detecting the presence of buried materials (waste) and of delineating the lateral extent of the landfill by identifying the electrical conductivity anomalies produced, for the most part, by the conductive waste fill. Using an EM31 device in the vertical-dipole configuration, at a height of 0.9 m above the ground, both quadrature and in-phase electromagnetic responses were collected over a 7-hectare area with elevation varying between 6 m and 2.8 m above sea level. After removing the measurements identified as data coming from any recognisable surface man-made features within the survey area or near its perimeter, the filtered quadrature response (expressed as apparent conductivity) ranged from 5.5 mS/m to about 188.6 mS/m. All values are beyond the low induction number (LIN) condition and valid for the classical EM31 mapping, thus requiring advanced data processing. To obtain undistorted, meaningful, and interpretable high-resolution maps, measured data have been processed to correct the bias, introduced by the nonlinearity of the device, as a function of height above ground and the topography. The comparative analysis of the apparent conductivity map, obtained by the properly processed EM31 data and some aerial photos that clearly documented the site history, has allowed unequivocal delineation of the landfill extent, in good agreement with the results obtained with other geophysical methods (not described in this paper) and with the ground truthing data provided by three boreholes, which were core-drilled at the end of the study at three locations selected on the basis of the apparent conductivity map

Intranasal Rapamycin Ameliorates Alzheimer-Like Cognitive Decline in a Mouse Model of Down Syndrome

Background: Down syndrome (DS) individuals, by the age of 40s, are at increased risk to develop Alzheimer-like dementia, with deposition in brain of senile plaques and neurofibrillary tangles. Our laboratory recently demonstrated the disturbance of PI3K/AKT/mTOR axis in DS brain, prior and after the development of Alzheimer Disease (AD). The aberrant modulation of the mTOR signalling in DS and AD age-related cognitive decline affects crucial neuronal pathways, including insulin signaling and autophagy, involved in pathology onset and progression. Within this context, the therapeutic use of mTOR-inhibitors may prevent/attenuate the neurodegenerative phenomena. By our work we aimed to rescue mTOR signalling in DS mice by a novel rapamycin intranasal administration protocol (InRapa) that maximizes brain delivery and reduce systemic side effects. Methods: Ts65Dn mice were administered with InRapa for 12 weeks, starting at 6 months of age demonstrating, at the end of the treatment by radial arms maze and novel object recognition testing, rescued cognition. Results: The analysis of mTOR signalling, after InRapa, demonstrated in Ts65Dn mice hippocampus the inhibition of mTOR (reduced to physiological levels), which led, through the rescue of autophagy and insulin signalling, to reduced APP levels, APP processing and APP metabolites production, as well as, to reduced tau hyperphosphorylation. In addition, a reduction of oxidative stress markers was also observed. Discussion: These findings demonstrate that chronic InRapa administration is able to exert a neuroprotective effect on Ts65Dn hippocampus by reducing AD pathological hallmarks and by restoring protein homeostasis, thus ultimately resulting in improved cognition. Results are discussed in term of a potential novel targeted therapeutic approach to reduce cognitive decline and AD-like neuropathology in DS individuals

Fishers’ Perception on the Interaction between Dolphins and Fishing Activities in Italian and Croatian Waters

Interactions between fishing and dolphins can be detrimental, since on one hand dolphins can be lethally entangled by nets and trawls, and on the other dolphins can predate fish caught by nets. For dolphins, this interaction can be dangerous as they can be wounded or accidentally killed; for fishers, the predation of their catch results in economic losses due to reduced quantity and/or quality of catches and damage to fishing gear. During July and November 2020, we surveyed the “dolphin–fisheries conflict” through compiling 209 fisher interviews from nine locations in Italy and Croatia. Fishers mentioned the common bottlenose dolphin (Tursiops truncatus) as the species primarily interacting with fishing, with the major issue being catch damage by predation. The interaction probability varied among gears and seasons, with some fishing activities (e.g., passive nets) more affected than others (e.g., bottom trawls), especially in terms of economic loss (1000–10,000 €/year on average). More than 70% of the fishers claimed that dolphin populations have increased over the last 10 years, in different degrees and based on different areas. Dolphin bycatch rates are generally low; however, 34.6% of respondents reported having captured at least one dolphin during their career. The fishers’ attitude towards acoustic deterrents (“pingers”) as a mitigation measure revealed that few of them were aware of these devices or were using them

The antidepressant fluoxetine acts on energy balance and leptin sensitivity via BDNF

Leptin and Brain Derived Neurotrophic Factor (BDNF) pathways are critical players in body weight homeostasis. Noninvasive treatments like environmental stimulation are able to increase response to leptin and induce BDNF expression in the brain. Emerging evidences point to the antidepressant selective serotonin reuptake inhibitor Fluoxetine (FLX) as a drug with effects similar to environmental stimulation. FLX is known to impact on body weight, with mechanisms yet to be elucidated. We herein asked whether FLX affects energy balance, the leptin system and BDNF function. Adult lean male mice chronically treated with FLX showed reduced weight gain, higher energy expenditure, increased sensitivity to acute leptin, increased hypothalamic BDNF expression, associated to changes in white adipose tissue expression typical of "brownization". In the Ntrk2tm1Ddg/J model, carrying a mutation in the BDNF receptor Tyrosine kinase B (TrkB), these effects are partially or totally reversed. Wild type obese mice treated with FLX showed reduced weight gain, increased energy output, and differently from untreated obese mice, a preserved acute response to leptin in terms of activation of the intracellular leptin transducer STAT3. In conclusion, FLX impacts on energy balance and induces leptin sensitivity and an intact TrkB function is required for these effects to take place

The ALLgorithMM: How to define the hemodilution of bone marrow samples in lymphoproliferative diseases

IntroductionMinimal residual disease (MRD) is commonly assessed in bone marrow (BM) aspirate. However, sample quality can impair the MRD measurement, leading to underestimated residual cells and to false negative results. To define a reliable and reproducible method for the assessment of BM hemodilution, several flow cytometry (FC) strategies for hemodilution evaluation have been compared. MethodsFor each BM sample, cells populations with a well-known distribution in BM and peripheral blood - e.g., mast cells (MC), immature (IG) and mature granulocytes (N) - have been studied by FC and quantified alongside the BM differential count. ResultsThe frequencies of cells' populations were correlated to the IG/N ratio, highlighting a mild correlation with MCs and erythroblasts (R=0.25 and R=0.38 respectively, with p-value=0.0006 and 0.0000052), whereas no significant correlation was found with B or T-cells. The mild correlation between IG/N, erythroblasts and MCs supported the combined use of these parameters to evaluate BM hemodilution, hence the optimization of the ALLgorithMM. Once validated, the ALLgorithMM was employed to evaluate the dilution status of BM samples in the context of MRD assessment. Overall, we found that 32% of FC and 52% of Next Generation Sequencing (NGS) analyses were MRD negative in samples resulted hemodiluted (HD) or at least mildly hemodiluted (mHD). ConclusionsThe high frequency of MRD-negative results in both HD and mHD samples implies the presence of possible false negative MRD measurements, impairing the correct assessment of patients' response to therapy and highlighs the importance to evaluate BM hemodilution

Antitumor Agents. 1. Synthesis, Biological Evaluation, and Molecular Modeling of 5H-Pyrido[3,2-a]phenoxazin-5-one, a Compound with Potent Antiproliferative Activity

The iminoquinone is an important moiety of a large number of antineoplastic drugs and plays a significant role in the nucleus of actinomycins, powerful, highly toxic, natural antibiotics that target DNA as intercalating agents. A series of polycyclic iminoquinonic compounds, 2-amino-3H-phenoxazin-3-one (1), 2-amino-1,9-diacetyl-3H-phenoxazin-3-one (2), 2-acetylamino- 3H-phenoxazin-3-one (3), 3H-phenoxazin-3-one (4), 5H-pyrido[3,2-a]phenoxazin-5-one (5), and 5H-pyrido[3,2-a]phenothiazin-5-one (6), strictly related to the actinomycin chromophore, were synthesized for developing new anticancer intercalating drugs. The antiproliferative activity of these compounds, evaluated against representative human liquid and solid neoplastic cell lines, showed that 5 and its isoster 6 were the most active compounds inhibiting cell proliferation in a submicromolar range. Compound 5 was also evaluated against KB subclones (KBMDR, KB7D, and KBV20C), which overexpress the MDR1/P-glycoprotein drug efflux pump responsible for drug resistance. All the above KB subclones did not show altered sensitivity to the antiproliferative activity of 5. UV-vis and 1H NMR spectroscopy experiments support the phenoxazinone 5/DNA binding. Molecular mechanics methods were used to build a three-dimensional model of the 5/[d(GAAGCTTC)]2 complex. Electrostatic interactions between the hydrogen of the positively charged pyridine nitrogen of 5 and the negatively charged oxygen atoms (O4¢ and O5¢) of the cytosine C5 residue together with stacking forces contribute to the high antiproliferative activity. The metal(II)-assisted synthesis procedure of 5 is described, and the formation mechanism is proposed